The objective of this project is to test the hypothesis that deregulation of genes for presenilins (PSs) and PS- interacting proteins contributes to Alzheimer's disease (AD) pathology. Missense-mutations in two homologous presenilin genes, PS1 and PS2, are the major cause of familial early onset Alzheimer's disease (AD). The role of these genes in the most common, the late-onsetAD, remains to be elucidated. PSs are involved in the proteolytic cleavage of several proteins, eg., amyloid precursor protein (APP), that is thought to be critical factor in molecular cascade mechanisms leading to AD. AD-associated mutations enhance this cleavage, while the abolishment of PS1 and PS2 activity inhibits the endoproteolysis of APP substrate. It is conceivable, then, that alteration of expression of PSs modulate AD pathology. Our preliminary data suggest hypoxia is an important factor in regulation of genes for PSs and PS- interacting proteins. The data strongly suggest that PS2 is highly inducible by hypoxia/hyperoxia conditions and that alteration of regulatory DNA elements (IRF2/CREBB) in PS2 increases the susceptibility of the promoter region to stress-regulated factors associated with AD. We also identified a novel family of presenilin-homologous proteins and highly regulated presenilin isoforms lacking functional domains. We hypothesize that activity of PSs is modulated by 1) transcriptional factors, including naturally-occurring DNA variations and stress-regulated DNA elements; and 2) post-transcriptional factors, including PS-like protein isoforms with putative dominant negative effects. The goal of this proposal is to elucidate molecular factors that regulate presenilins on transcriptional and posttranslational levels. We will 1) identify novel-molecular-genetic factors (gene haplotypes) contributing to AD; 2) isolate novel transcription and splicing enhancer/repressor elements that dynamically regulate PSs; 3) elucidate the role of newly identified PS-homologous proteins in modulation of processing or expression of PS proteins. We present a convergent approach with which the genetic analysis of AD patients will be accompanied by phylogenic and functional analysis of the gene variations in vitro. To achieve these goals, methods of genetic association analysis, molecular assays for quantitative evaluation of promoter activities, and protein processing in mammalian cells, cellular assays for exon trapping and PS-mediated proteolysis will be undertaken. The accomplishment of these goals will clarify molecular-genetic and epigenetic mechanisms of AD. These studies will contribute to identification of molecular factors regulating the primary causes of AD, and ultimately, to the development of new strategies for prevention and rational therapy of Alzheimer's disease.